Diagnosis of cystic lung diseases: a position statement from the UK Cystic Lung Disease Rare Disease Collaborative Network.

BACKGROUND: Rare cystic lung diseases are increasingly recognised due the wider application of CT scanning making cystic lung disease management a growing part of respiratory care. Cystic lung diseases tend to have extrapulmonary features that can both be diagnostic but also require surveillance and treatment in their own right. As some of these diseases now have specific treatments, making a precise diagnosis is crucial. While Langerhans cell histiocytosis, Birt-Hogg-Dubé syndrome, lymphoid interstitial pneumonia and lymphangioleiomyomatosis are becoming relatively well-known diseases to respiratory physicians, a targeted and thorough workup improves diagnostic accuracy and may suggest other ultrarare diseases such as light chain deposition disease, cystic pulmonary amyloidosis, low-grade metastatic neoplasms or infections. In many cases, diagnostic information is overlooked leaving uncertainty over the disease course and treatments. AIMS: This position statement from the Rare Disease Collaborative Network for cystic lung diseases will review how clinical, radiological and physiological features can be used to differentiate between these diseases. NARRATIVE: We highlight that in many cases a multidisciplinary diagnosis can be made without the need for lung biopsy and discuss where tissue sampling is necessary when non-invasive methods leave diagnostic doubt. We suggest an initial workup focusing on points in the history which identify key disease features, underlying systemic and familial diseases and a clinical examination to search for connective tissue disease and features of genetic causes of lung cysts. All patients should have a CT of the thorax and abdomen to characterise the pattern and burden of lung cysts and extrapulmonary features and also spirometry, gas transfer and a 6 min walk test. Discussion with a rare cystic lung disease centre is suggested before a surgical biopsy is undertaken. CONCLUSIONS: We suggest that this focused workup should be performed in all people with multiple lung cysts and would streamline referral pathways, help guide early treatment, management decisions, improve patient experience and reduce overall care costs. It could also potentially catalyse a national research database to describe these less well-understood and unidentified diseases, categorise disease phenotypes and outcomes, potentially leading to better prognostic data and generating a stronger platform to understand specific disease biology.

Asthma and COPD: A Focus on ß-Agonists - Past, Present and Future.

Asthma has been recognised as a respiratory disorder for millennia and the focus of targeted drug development for the last 120 years. Asthma is one of the most common chronic non-communicable diseases worldwide. Chronic obstructive pulmonary disease (COPD), a leading cause of morbidity and mortality worldwide, is caused by exposure to tobacco smoke and other noxious particles and exerts a substantial economic and social burden. This chapter reviews the development of the treatments of asthma and COPD particularly focussing on the ß-agonists, from the isolation of adrenaline, through the development of generations of short- and long-acting ß-agonists. It reviews asthma death epidemics, considers the intrinsic efficacy of clinical compounds, and charts the improvement in selectivity and duration of action that has led to our current medications. Important ß2-agonist compounds no longer used are considered, including some with additional properties, and how the different pharmacological properties of current ß2-agonists underpin their different places in treatment guidelines. Finally, it concludes with a look forward to future developments that could improve the ß-agonists still further, including extending their availability to areas of the world with less readily accessible healthcare.

The role of inflammation in anxiety and depression in the European U-BIOPRED asthma cohorts.

BACKGROUND: Growing evidence indicates high comorbid anxiety and depression in patients with asthma. However, the mechanisms underlying this comorbid condition remain unclear. The aim of this study was to investigate the role of inflammation in comorbid anxiety and depression in three asthma patient cohorts of the Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes (U-BIOPRED) project. METHODS: U-BIOPRED was conducted by a European Union consortium of 16 academic institutions in 11 European countries. A subset dataset from subjects with valid anxiety and depression measures and a large blood biomarker dataset were analysed, including 198 non-smoking patients with severe asthma (SAn), 65 smoking patients with severe asthma (SAs), 61 non-smoking patients with mild-to-moderate asthma (MMA), and 20 healthy non-smokers (HC). The Hospital Anxiety and Depression Scale was used to measure anxiety and depression and a series of inflammatory markers were analysed by the SomaScan v3 platform (SomaLogic, Boulder, Colo). ANOVA and the Kruskal-Wallis test were used for multiple-group comparisons as appropriate. RESULTS: There were significant group effects on anxiety and depression among the four cohort groups (p < 0.05). Anxiety and depression of SAn and SAs groups were significantly higher than that of MMA and HC groups (p < 0.05. There were significant differences in serum IL6, MCP1, CCL18, CCL17, IL8, and Eotaxin among the four groups (p < 0.05). Depression was significantly associated with IL6, MCP1, CCL18 level, and CCL17; whereas anxiety was associated with CCL17 only (p < 0.05). CONCLUSIONS: The current study suggests that severe asthma patients are associated with higher levels of anxiety and depression, and inflammatory responses may underlie this comorbid condition.

Effect of implementing the NEWS2 escalation protocol in a large acute NHS trust: a retrospective cohort analysis of mortality, workload and ability of early warning score to predict death within 24 hours.

OBJECTIVES: To describe the inpatient population, establish patterns in admission and mortality over a 4-year period in different cohorts and assess the prognostic ability and workload implications of introducing the National Early Warning Score 2 (NEWS2) and associated escalation protocol. DESIGN: Retrospective cohort analyses of medical and surgical inpatient admissions. SETTING: Large teaching hospital with tertiary inpatient care and a major trauma centre employing an electronic observations platform, initially with a local early warning score, followed by NEWS2 introduction in June 2019. PARTICIPANTS: 332 682 adult patients were admitted between 1 January 2016 and 31 December 2019. OUTCOME MEASURES: Mortality, workload and ability of early warning score to predict death within 24 hours. RESULTS: Admissions rose by 19% from 76 055 in 2016 to 90 587 in 2019. Total bed days rose by 10% from 433 382 to 477 485. Mortality fell from 3.7% to 3.1% and was significantly lower in patients discharged from a surgical specialty, 1.0%-1.2% (p<0.001). Total observations recorded increased by 14% from 1 976 872 in 2016 to 2 249 118 in 2019. 65% of observations were attributable to patients under medical specialties, 34% to patients under surgical specialties. Recorded escalations to the registrar were stable from January 2016 to May 2019 but trebled following the introduction of NEWS2 in June 2019. CONCLUSIONS: There was an increase in hospital inpatient activity between 2016 and 2019, associated with a reduction in mortality and percentage of observations calculated as reaching threshold NEWS2 score of 7 for escalation to the registrar. The introduction of the NEWS2, with a higher sensitivity and lower specificity, when allied to its escalation protocol, was associated with a significant increase in actual recorded escalations to the registrar. This was more marked in the surgical population and would support refining threshold scores based on admission characteristics when developing the next iteration of NEWS.

Impact of introducing procalcitonin testing on antibiotic usage in acute NHS hospitals during the first wave of COVID-19 in the UK: a controlled interrupted time series analysis of organization-level data.

BACKGROUND: Blood biomarkers have the potential to help identify COVID-19 patients with bacterial coinfection in whom antibiotics are indicated. During the COVID-19 pandemic, procalcitonin testing was widely introduced at hospitals in the UK to guide antibiotic prescribing. We have determined the impact of this on hospital-level antibiotic consumption. METHODS: We conducted a retrospective, controlled interrupted time series analysis of organization-level data describing antibiotic dispensing, hospital activity and procalcitonin testing for acute hospitals/hospital trusts in England and Wales during the first wave of COVID-19 (24 February to 5 July 2020). RESULTS: In the main analysis of 105 hospitals in England, introduction of procalcitonin testing in emergency departments/acute medical admission units was associated with a statistically significant decrease in total antibiotic use of -1.08 (95% CI: -1.81 to -0.36) DDDs of antibiotic per admission per week per trust. This effect was then lost at a rate of 0.05 (95% CI: 0.02-0.08) DDDs per admission per week. Similar results were found specifically for first-line antibiotics for community-acquired pneumonia and for COVID-19 admissions rather than all admissions. Introduction of procalcitonin in the ICU setting was not associated with any significant change in antibiotic use. CONCLUSIONS: At hospitals where procalcitonin testing was introduced in emergency departments/acute medical units this was associated with an initial, but unsustained, reduction in antibiotic use. Further research should establish the patient-level impact of procalcitonin testing in this population and understand its potential for clinical effectiveness.

Airway Elastin is increased in severe asthma and relates to proximal wall area: histological and computed tomography findings from the U-BIOPRED severe asthma study.

BACKGROUND: Airway remodelling, which may include goblet cell hyperplasia / hypertrophy, changes in epithelial integrity, accumulation of extracellular matrix components, smooth muscle hypertrophy and thickening of the lamina reticularis, is a feature of severe asthma and contributes to the clinical phenotype. OBJECTIVE: Within the U-BIOPRED severe asthma study, we have assessed histological elements of airway remodelling and their relationship to computed tomography (CT) measures of proximal airway dimensions. METHODS: Bronchial biopsies were collected from two severe asthma groups, one non-smoker (SAn, n = 28) and one current/ex-smoker (SAs/ex, n = 13), and a mild-moderate asthma group (MMA, n = 28) classified and treated according to GINA guidelines, plus a healthy control group (HC, n = 33). Movat's pentachrome technique was used to identify mucin, elastin and total collagen in these biopsies. The number of goblet cells (mucin+) was counted as a percentage of the total number of epithelial cells and the percentage mucin epithelial area measured. The percentage area of elastic fibres and total collagen within the submucosa was also measured, and the morphology of the elastic fibres classified. Participants in the asthma groups also had a CT scan to assess large airway morphometry. RESULTS: The submucosal tissue elastin percentage was higher in both severe asthma groups (16.1% SAn, 18.9% SAs/ex) compared with the HC (9.7%) but did not differ between asthma groups. There was a positive relationship between elastin and airway wall area measured by CT (n = 18-20, rho=0.544, p = 0.024), which also related to an increase in elastic fibres with a thickened lamellar morphological appearance. Mucin epithelial area and total collagen were not different between the four groups. Due to small numbers of suitable CT scans, it was not feasible to compare airway morphometry between the asthma groups. CONCLUSION: These findings identify a link between extent of elastin deposition and airway wall thickening in severe asthma.

Urinary Leukotriene E4 and Prostaglandin D2 Metabolites Increase in Adult and Childhood Severe Asthma Characterized by Type 2 Inflammation. A Clinical Observational Study.

Rationale: New approaches are needed to guide personalized treatment of asthma.Objectives: To test if urinary eicosanoid metabolites can direct asthma phenotyping.Methods: Urinary metabolites of prostaglandins (PGs), cysteinyl leukotrienes (CysLTs), and isoprostanes were quantified in the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Diseases Outcomes) study including 86 adults with mild-to-moderate asthma (MMA), 411 with severe asthma (SA), and 100 healthy control participants. Validation was performed internally in 302 participants with SA followed up after 12-18 months and externally in 95 adolescents with asthma.Measurement and Main Results: Metabolite concentrations in healthy control participants were unrelated to age, body mass index, and sex, except for the PGE2 pathway. Eicosanoid concentrations were generally greater in participants with MMA relative to healthy control participants, with further elevations in participants with SA. However, PGE2 metabolite concentrations were either the same or lower in male nonsmokers with asthma than in healthy control participants. Metabolite concentrations were unchanged in those with asthma who adhered to oral corticosteroid treatment as documented by urinary prednisolone detection, whereas those with SA treated with omalizumab had lower concentrations of LTE4 and the PGD2 metabolite 2,3-dinor-11ß-PGF2α. High concentrations of LTE4 and PGD2 metabolites were associated with lower lung function and increased amounts of exhaled nitric oxide and eosinophil markers in blood, sputum, and urine in U-BIOPRED participants and in adolescents with asthma. These type 2 (T2) asthma associations were reproduced in the follow-up visit of the U-BIOPRED study and were found to be as sensitive to detect T2 inflammation as the established biomarkers.Conclusions: Monitoring of urinary eicosanoids can identify T2 asthma and introduces a new noninvasive approach for molecular phenotyping of adult and adolescent asthma.Clinical trial registered with www.clinicaltrials.gov (NCT01976767).

Hospital doctors' attitudes to brief educational messages that aim to modify diagnostic test requests: a qualitative study.

BACKGROUND: Avoidable use of diagnostic tests can both harm patients and increase the cost of healthcare. Nudge-type educational interventions have potential to modify clinician behaviour while respecting clinical autonomy and responsibility, but there is little evidence how this approach may be best used in a healthcare setting. This study aims to explore attitudes of hospital doctors to two nudge-type messages: one concerning potential future cancer risk after receiving a CT scan, another about the financial costs of blood tests. METHODS: We added two brief educational messages to diagnostic test results in a UK hospital for one year. One message on the associated long-term potential cancer risk from ionising radiation imaging to CT scan reports, and a second on the financial costs incurred to common blood test results. We conducted a qualitative study involving telephone interviews with doctors working at the hospital to identify themes explaining their response to the intervention. RESULTS: Twenty eight doctors were interviewed. Themes showed doctors found the intervention to be highly acceptable, as the group had a high awareness of the need to prevent harm and optimise use of finite resources, and most found the nudge-type approach to be inoffensive and harmless. However, the messages were not seen as personally relevant because doctors felt they were already relatively conservative in their use of tests. Cancer risk was important in decision-making but was not considered to represent new knowledge to doctors. Conversely, financial costs were considered to be novel information that was unimportant in decision-making. Defensive medicine was commonly cited as a barrier to individual behaviour change. The educational cancer risk message on CT scan reports increased doctors' confidence to challenge decisions and explain risks to patients and there were some modifications in clinical practice prompted by the financial cost message. CONCLUSION: The nudge-type approach to target avoidable use of tests was acceptable to hospital doctors but there were barriers to behaviour change. There was evidence doctors perceived this cheap and light-touch method can contribute to culture change and form a foundation for more comprehensive educational efforts to modify behaviour in a healthcare environment.

Evaluation of the impact of a brief educational message on clinicians' awareness of risks of ionising-radiation exposure in imaging investigations: a pilot pre-post intervention study.

BACKGROUND: In the context of increasing availability of computed tomography (CT) scans, judicious use of ionising radiation is a priority to minimise the risk of future health problems. Hence, education of clinicians on the risks and benefits of CT scans in the management of patients is important. METHODS: An educational message about the associated lifetime cancer risk of a CT scan was added to all CT scan reports at a busy acute teaching hospital in the UK. An online multiple choice survey was completed by doctors before and after the intervention, assessing education and knowledge of the risks involved with exposure to ionising radiation. RESULTS: Of 546 doctors contacted at baseline, 170 (31%) responded. Over a third (35%) of respondents had received no formal education on the risks of exposure to ionising radiation. Over a quarter (27%) underestimated (selected 1 in 30,000 or negligible lifetime cancer risk) the risk associated with a chest, abdomen and pelvis CT scan for a 20 year old female. Following exposure to the intervention for 1 year there was a statistically significant improvement in plausible estimates of risk from 68.3 to 82.2% of respondents (p < 0.001). There was no change in the proportion of doctors correctly identifying imaging modalities that do or do not involve ionising radiation. CONCLUSIONS: Training on the longterm risks associated with diagnostic radiation exposure is inadequate among hospital doctors. Exposure to a simple non-directional educational message for 1 year improved doctors' awareness of risks associated with CT scans. This demonstrates the potential of the approach to improve knowledge that could improve clinical practice. This approach is easily deliverable and may have applications in other areas of clinical medicine. The wider and longer term impact on radiation awareness is unknown, however, and there may be a need for regular mandatory training in the risks of radiation exposure.

Airway epithelial cell isolation techniques affect DNA methylation profiles with consequences for analysis of asthma related perturbations to DNA methylation.

The airway epithelium forms the interface between the inhaled environment and the lung. The airway epithelium is dysfunctional in asthma and epigenetic mechanisms are considered a contributory factor. We hypothesised that the DNA methylation profiles of cultured primary airway epithelial cells (AECs) would differ between cells isolated from individuals with asthma (n = 17) versus those without asthma (n = 16). AECs were isolated from patients by two different isolation techniques; pronase digestion (9 non-asthmatic, 8 asthmatic) and bronchial brushings (7 non-asthmatic and 9 asthmatic). DNA methylation was assessed using an Illumina Infinium HumanMethylation450 BeadChip array. DNA methylation of AECs clustered by isolation technique and linear regression identified 111 CpG sites differentially methylated between isolation techniques in healthy individuals. As a consequence, the effect of asthmatic status on DNA methylation was assessed within AEC samples isolated using the same technique. In pronase isolated AECs, 15 DNA regions were differentially methylated between asthmatics and non-asthmatics. In bronchial brush isolated AECs, 849 differentially methylated DNA regions were identified with no overlap to pronase regions. In conclusion, regardless of cell isolation technique, differential DNA methylation was associated with asthmatic status in AECs, providing further evidence for aberrant DNA methylation as a signature of epithelial dysfunction in asthma.

Short and Long term predictions of Hospital emergency department attendances.

OBJECTIVE: Emergency departments in the United Kingdom (UK) experience significant difficulties in achieving the 95% NHS access standard due to unforeseen variations in patient flow. In order to maximize efficiency and minimize clinical risk, better forecasting of patient demand is necessary. The objective is therefore to create a tool that accurately predicts attendance at emergency departments to support optimal planning of human and physical resources. METHODS: Historical attendance data between Jan-2011 - December-2015 from four hospitals were used as a training set to develop and validate a forecasting model. To handle weekday variations, the data was first segmented into each weekday time series and a separate model for each weekday was performed. Seasonality testing was performed, followed by Box-Cox transformations. A modified heuristics based on a fuzzy time series model was then developed and compared with autoregressive integrated moving average and neural networks models using Harvey, Leybourne and Newbold (HLN) test. The time series models were tested in four emergency department sites to assess forecasting accuracy using the root mean square error and mean absolute percentage error. The models were tested for (i) short term prediction (four weeks ahead), using weekday time series; and (ii) long term predictions (four months ahead) using monthly time series. RESULTS: Data analysis revealed that presentations to emergency department and subsequent admissions to hospital were not a purely random process and therefore could be predicted with acceptable accuracy. Prediction accuracy improved as the forecast time intervals became wider (from daily to monthly). For each weekday time series modelling using fuzzy time series, for forecasting daily admissions, the mean absolute percentage error ranged from 2.63% to 4.72% while for monthly time series mean absolute percentage error varied from 2.01%-2.81%. For weekday time series, the mean absolute percentage error for autoregressive integrated moving average and neural network forecasting models ranged from 6.25% to 7.47% and 6.04%-7.42% respectively. The proposed fuzzy time series model proved to have statistically significant performance using Harvey, Leybourne and Newbold (HLN) test. This was explained by variations in attendances in different sites and weekdays. CONCLUSIONS: This paper described a heuristic-based fuzzy logic model for predicting emergency department attendances which could help resource allocation and reduce pressure on busy hospitals. Valid and reproducible prediction tools could be generated from these hospital data. The methodology had an acceptable accuracy over a relatively short time period, and could be used to assist better bed management, staffing and elective surgery scheduling. When compared to other prediction models usually applied for emergency department attendances prediction, the proposed heuristic model had better accuracy.

Evaluation of a nudge intervention providing simple feedback to clinicians of the consequence of radiation exposure on demand for computed tomography: a controlled study.

Computed tomography (CT) is readily available in developed countries. As one of the side effects includes an increased risk of cancer, interventions that may encourage more judicious use of CT are important. Behavioural economics theory includes the use of nudges that aim to help more informed decisions to be made, although these have been rarely used in hospitals to date. We aimed to evaluate the impact of a simple educational message appended to the CT report on subsequent numbers of CT completed using a controlled interrupted time series design based in two teaching hospitals in the UK. The intervention was the addition of a non-directional educational message on the risk of ionising radiation to all CT reports. There was a statistically significant reduction in the number of CT requested in the intervention hospital compared to the control hospital (-4.6%, 95% confidence intervals -7.4 to -1.7, p=0.002) in the 12 months after the intervention was implemented. We conclude that a simple, non-directional nudge intervention has the capacity to modify clinician use of CT. This approach is cheap, and has potential in helping support doctors make informed decisions.

IL-17-high asthma with features of a psoriasis immunophenotype.

BACKGROUND: The role of IL-17 immunity is well established in patients with inflammatory diseases, such as psoriasis and inflammatory bowel disease, but not in asthmatic patients, in whom further study is required. OBJECTIVE: We sought to undertake a deep phenotyping study of asthmatic patients with upregulated IL-17 immunity. METHODS: Whole-genome transcriptomic analysis was performed by using epithelial brushings, bronchial biopsy specimens (91 asthmatic patients and 46 healthy control subjects), and whole blood samples (n = 498) from the Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes (U-BIOPRED) cohort. Gene signatures induced in vitro by IL-17 and IL-13 in bronchial epithelial cells were used to identify patients with IL-17-high and IL-13-high asthma phenotypes. RESULTS: Twenty-two of 91 patients were identified with IL-17, and 9 patients were identified with IL-13 gene signatures. The patients with IL-17-high asthma were characterized by risk of frequent exacerbations, airway (sputum and mucosal) neutrophilia, decreased lung microbiota diversity, and urinary biomarker evidence of activation of the thromboxane B2 pathway. In pathway analysis the differentially expressed genes in patients with IL-17-high asthma were shared with those reported as altered in psoriasis lesions and included genes regulating epithelial barrier function and defense mechanisms, such as IL1B, IL6, IL8, and ß-defensin. CONCLUSION: The IL-17-high asthma phenotype, characterized by bronchial epithelial dysfunction and upregulated antimicrobial and inflammatory response, resembles the immunophenotype of psoriasis, including activation of the thromboxane B2 pathway, which should be considered a biomarker for this phenotype in further studies, including clinical trials targeting IL-17.

Idiopathic chronic productive cough and response to open-label macrolide therapy: An observational study.

BACKGROUND AND OBJECTIVE: Adult patients with chronic productive cough of unknown cause are commonly seen in respiratory clinics. We have previously described a subgroup of these patients who have a short-lived response to standard antibiotic treatment but a prolonged response to 3 months of low-dose azithromycin therapy. METHODS: This observational study describes the physiological, radiological and pathological features of this patient cohort along with their response to a 12-week open-label trial of 250 mg azithromycin thrice weekly. RESULTS: A total of 30 subjects with a mean age of 57 were recruited. The majority demonstrated airway dilatation on high-resolution computed tomography (HRCT) scan without evidence of established bronchiectasis (n = 21) and non-specific chronic inflammatory changes on bronchial biopsy (n = 15/17). Twenty-nine subjects completed 3 months of azithromycin with a significant improvement in median Leicester Cough Questionnaire (LCQ) score (-6.3 points, P < 0.00001), reduction in median 24-h sputum volume (-5.8 mL, P = 0.0003) and improvement in sputum colour (P = 0.003). Patients responsive to azithromycin (n = 22) demonstrated neutrophilic or paucigranulocytic airway inflammation, whereas five subjects with eosinophilic airways inflammation did not respond symptomatically to azithromycin. CONCLUSION: We describe a cohort of patients with chronic productive cough not adequately described by existing disease labels whose symptoms responded well to low-dose azithromycin. Many of the features are similar to the paediatric condition protracted bacterial bronchitis.

Contribution of airway eosinophils in airway wall remodeling in asthma: Role of MMP-10 and MET.

BACKGROUND: Eosinophils play an important role in the pathophysiology of asthma being implicated in airway epithelial damage and airway wall remodeling. We determined the genes associated with airway remodeling and eosinophilic inflammation in patients with asthma. METHODS: We analyzed the transcriptomic data from bronchial biopsies of 81 patients with moderate-to-severe asthma of the U-BIOPRED cohort. Expression profiling was performed using Affymetrix arrays on total RNA. Transcription binding site analysis used the PRIMA algorithm. Localization of proteins was by immunohistochemistry. RESULTS: Using stringent false discovery rate analysis, MMP-10 and MET were significantly overexpressed in biopsies with high mucosal eosinophils (HE) compared to low mucosal eosinophil (LE) numbers. Immunohistochemical analysis confirmed increased expression of MMP-10 and MET in bronchial epithelial cells and in subepithelial inflammatory and resident cells in asthmatic biopsies. Using less-stringent conditions (raw P-value < 0.05, log2 fold change > 0.5), we defined a 73-gene set characteristic of the HE compared to the LE group. Thirty-three of 73 genes drove the pathway annotation that included extracellular matrix (ECM) organization, mast cell activation, CC-chemokine receptor binding, circulating immunoglobulin complex, serine protease inhibitors, and microtubule bundle formation pathways. Genes including MET and MMP10 involved in ECM organization correlated positively with submucosal thickness. Transcription factor binding site analysis identified two transcription factors, ETS-1 and SOX family proteins, that showed positive correlation with MMP10 and MET expression. CONCLUSION: Pathways of airway remodeling and cellular inflammation are associated with submucosal eosinophilia. MET and MMP-10 likely play an important role in these processes.

Epithelial IL-6 trans-signaling defines a new asthma phenotype with increased airway inflammation.

BACKGROUND: Although several studies link high levels of IL-6 and soluble IL-6 receptor (sIL-6R) to asthma severity and decreased lung function, the role of IL-6 trans-signaling (IL-6TS) in asthmatic patients is unclear. OBJECTIVE: We sought to explore the association between epithelial IL-6TS pathway activation and molecular and clinical phenotypes in asthmatic patients. METHODS: An IL-6TS gene signature obtained from air-liquid interface cultures of human bronchial epithelial cells stimulated with IL-6 and sIL-6R was used to stratify lung epithelial transcriptomic data (Unbiased Biomarkers in Prediction of Respiratory Disease Outcomes [U-BIOPRED] cohorts) by means of hierarchical clustering. IL-6TS-specific protein markers were used to stratify sputum biomarker data (Wessex cohort). Molecular phenotyping was based on transcriptional profiling of epithelial brushings, pathway analysis, and immunohistochemical analysis of bronchial biopsy specimens. RESULTS: Activation of IL-6TS in air-liquid interface cultures reduced epithelial integrity and induced a specific gene signature enriched in genes associated with airway remodeling. The IL-6TS signature identified a subset of patients with IL-6TS-high asthma with increased epithelial expression of IL-6TS-inducible genes in the absence of systemic inflammation. The IL-6TS-high subset had an overrepresentation of frequent exacerbators, blood eosinophilia, and submucosal infiltration of T cells and macrophages. In bronchial brushings Toll-like receptor pathway genes were upregulated, whereas expression of cell junction genes was reduced. Sputum sIL-6R and IL-6 levels correlated with sputum markers of remodeling and innate immune activation, in particular YKL-40, matrix metalloproteinase 3, macrophage inflammatory protein 1ß, IL-8, and IL-1ß. CONCLUSIONS: Local lung epithelial IL-6TS activation in the absence of type 2 airway inflammation defines a novel subset of asthmatic patients and might drive airway inflammation and epithelial dysfunction in these patients.

Enhanced oxidative stress in smoking and ex-smoking severe asthma in the U-BIOPRED cohort.

Oxidative stress is believed to be a major driver of inflammation in smoking asthmatics. The U-BIOPRED project recruited a cohort of Severe Asthma smokers/ex-smokers (SAs/ex) and non-smokers (SAn) with extensive clinical and biomarker information enabling characterization of these subjects. We investigated oxidative stress in severe asthma subjects by analysing urinary 8-iso-PGF2α and the mRNA-expression of the main pro-oxidant (NOX2; NOSs) and anti-oxidant (SODs; CAT; GPX1) enzymes in the airways of SAs/ex and SAn. All the severe asthma U-BIOPRED subjects were further divided into current smokers with severe asthma (CSA), ex-smokers with severe asthma (ESA) and non-smokers with severe asthma (NSA) to deepen the effect of active smoking. Clinical data, urine and sputum were obtained from severe asthma subjects. A bronchoscopy to obtain bronchial biopsy and brushing was performed in a subset of subjects. The main clinical data were analysed for each subset of subjects (urine-8-iso-PGF2α; IS-transcriptomics; BB-transcriptomics; BBr-transcriptomics). Urinary 8-iso-PGF2α was quantified using mass spectrometry. Sputum, bronchial biopsy and bronchial brushing were processed for mRNA expression microarray analysis. Urinary 8-iso-PGF2α was increased in SAs/ex, median (IQR) = 31.7 (24.5-44.7) ng/mmol creatinine, compared to SAn, median (IQR) = 26.6 (19.6-36.6) ng/mmol creatinine (p< 0.001), and in CSA, median (IQR) = 34.25 (24.4-47.7), vs. ESA, median (IQR) = 29.4 (22.3-40.5), and NSA, median (IQR) = 26.5 (19.6-16.6) ng/mmol creatinine (p = 0.004). Sputum mRNA expression of NOX2 was increased in SAs/ex compared to SAn (probe sets 203922_PM_s_at fold-change = 1.05 p = 0.006; 203923_PM_s_at fold-change = 1.06, p = 0.003; 233538_PM_s_at fold-change = 1.06, p = 0.014). The mRNA expression of antioxidant enzymes were similar between the two severe asthma cohorts in all airway samples. NOS2 mRNA expression was decreased in bronchial brushing of SAs/ex compared to SAn (fold-change = -1.10; p = 0.029). NOS2 mRNA expression in bronchial brushing correlated with FeNO (Kendal's Tau = 0.535; p< 0.001). From clinical and inflammatory analysis, FeNO was lower in CSA than in ESA in all the analysed subject subsets (p< 0.01) indicating an effect of active smoking. Results about FeNO suggest its clinical limitation, as inflammation biomarker, in severe asthma active smokers. These data provide evidence of greater systemic oxidative stress in severe asthma smokers as reflected by a significant changes of NOX2 mRNA expression in the airways, together with elevated urinary 8-iso-PGF2α in the smokers/ex-smokers group. Trial registration ClinicalTrials.gov-Identifier: NCT01976767.